Cell processing apparatus, sample preparation apparatus, and cell analyzer

ABSTRACT

A cell processing apparatus includes a storage container that contains liquid L including a biological sample; a filter that prevents a first cell C 1  in the biological sample from passing and allows a second cell C 2  having a smaller diameter than that of the first cell C 1  to pass; and a filtration cylinder for separating, in the storage container and via the filter, the liquid L into a first liquid L 1  mainly including the first cell C 1  and a second liquid L 2  mainly including the second cell C 2 . A measurement target cell filtered by the filter among other cells can be easily collected.

RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.12/892,579 filed on Sep. 28, 2010 and issued on Apr. 9, 2013 as U.S.Pat. No. 8,415,145, which is a continuation of PCT/JP2009/056071 filedon Mar. 26, 2009, which claims priority to Japanese Application Nos.JP2008-089863 and JP2008-089868 both filed on Mar. 31, 2008. The entirecontents of these applications are incorporated herein by reference.

BACKGROUND OF THE INVENTION

1. Technical Field

The present invention relates to a cell processing apparatus, a samplepreparation apparatus, and a cell analyzer.

Specifically, the present invention relates to a technique for easilycollecting a measurement target cell that is discriminated by a filterfrom other cells.

2. Background Art

Conventionally, as a cell analyzer for analyzing cells included in abiological sample extracted from a living body, there has been known acell analyzer in which epidermal cells of a cervix included in a sampleextracted from the cervix of a subject are measured by a flow cytometerto perform the screening of cancer cells and atypical cells (e.g., seethe pamphlet of International Publication No. 2006/103920).

In the case of the cell analyzer disclosed in the above pamphlet, anepidermal cell of a cervix is assumed as a measurement target. A sampleextracted from a cervix includes not only an epidermal cell but alsocells such as red blood cells and white blood cells. If the sample ismeasured without any processing, the measurement result is influenced bythe cells such as red blood cells or the white blood cells. This mayprevent an accurate screening of cancer and atypical cells.

Thus, there has been required a technique for discriminating ameasurement target cell and cells other than the measurement target cellto easily collect only the measurement target cell.

As a technique for performing the above collection, a cellseparation/collection apparatus has been known for example in which cellsuspension liquid including blood cells is allowed to flow on a filterand a lymphocyte as a measurement target cell is captured by the filter.Then, liquid including other cells such as red blood cells or whiteblood cells is discharged through the filter and collection liquid isthen supplied to the filter to collect lymphocytes captured by thefilter (for example, see U.S. Pat. No. 6,312,950).

However, in the case of the above cell separation/collection apparatusdisclosed in U.S. Pat. No. 6,312,950, only a lymphocyte is captured byan upper face of the filter. However, liquid that included thelymphocyte is discharged to the outside through the filter, thusrequiring an operation to additionally separate the lymphocyte attachedto the filter to collect the lymphocyte.

Thus, in the case of the cell separation/collection apparatus disclosedin U.S. Pat. No. 6,312,950, a disadvantage is caused where theseparation of a lymphocyte from a filter requires an operation to changea cavity rate of the filter to a cavity rate different from that used tocapture a lymphocyte and an operation to supply collection liquid to thefilter, thus requiring a complicated mechanism.

It is noted that, a technique also has been known by which, when ameasurement target cell has a smaller diameter than those of othercells, sample liquid including a measurement target cell is sent to afilter, and the liquid including the measurement target cell is allowedto pass through the filter and is collected, and other cells arecaptured by the filter and are discharged.

However, this technique allows the measurement target cell to passthrough the filter. Thus, this technique cannot be used to a case as inan epidermal cell where a measurement target cell has a larger diameterthan those of other cells.

SUMMARY OF THE INVENTION

The scope of the present invention is defined solely by the appendedclaims, and is not affected to any degree by the statements within thissummary.

A first aspect of the present invention is a sample preparationapparatus comprising: a storage container that can contain liquidincluding a biological sample; a filter that prevents a first cell inthe biological sample from passing therethrough and that allows a secondcell having a smaller diameter than that of the first cell to passtherethrough; a liquid separation section for separating, in the storagecontainer and via the filter, the liquid into a first liquid mainlyincluding the first cell and a second liquid mainly including the secondcell, so that the first liquid is below the filter while the secondliquid is above the filter; a liquid acquisition section for acquiringthe first liquid separated so as to be below the filter by the liquidseparation section; and a sample preparation section for preparing ameasurement sample from the first liquid acquired by the liquidacquisition section and a predetermined reagent.

A second aspect of the present invention is a cell analyzer comprising:a storage container that can contain liquid including a biologicalsample; a filter that prevents a first cell in the biological samplefrom passing therethrough and that allows a second cell having a smallerdiameter than that of the first cell to pass therethrough; a liquidseparation section for separating, in the storage container and via thefilter, the liquid into a first liquid mainly including the first celland a second liquid mainly including the second cell, so that the firstliquid is below the filter while the second liquid is above the filter;a liquid acquisition section for acquiring the first liquid separated soas to be below the filter by the liquid separation section; a detectionsection for detecting the first cell from the first liquid acquired bythe liquid acquisition section; and an analysis section for analyzingthe first cell based on a detection result by the detection section.

A third aspect of the present invention is a cell processing apparatuscomprising: a storage container that can contain liquid including abiological sample; a filter that prevents a first cell in the biologicalsample from passing therethrough and that allows a second cell having asmaller diameter than that of the first cell to pass therethrough; aliquid separation section for separating, in the storage container andvia the filter, the liquid into a first liquid mainly including thefirst cell and a second liquid mainly including the second cell, so thatthe first liquid is below the filter while the second liquid is abovethe filter; and a liquid acquisition section for acquiring the firstliquid separated so as to be below the filter by the liquid separationsection.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a perspective view illustrating a cell analyzer according tothe first embodiment;

FIG. 2 is a block diagram illustrating the internal configuration of ameasurement apparatus;

FIG. 3 is a block diagram illustrating the internal configuration of asample preparation apparatus;

FIG. 4 is a block diagram illustrating the internal configuration of adata processing apparatus;

FIG. 5 is a function block diagram of a flow cytometer configuring amain detecting section;

FIG. 6 is a side view illustrating an optical system of the flowcytometer;

FIG. 7 is a fluid circuit diagram of a preparation device section;

FIG. 8 is an expanded cross-sectional view illustrating a more specificconfiguration of a discrimination/substitution section;

FIG. 9(a)-9(c) are explanatory diagrams showing the filtration action ofthe discrimination/substitution section;

FIG. 10 is a flowchart illustrating the processings performed by therespective control sections of the cell analyzer;

FIG. 11 is a flowchart illustrating the processings performed by therespective control sections of the cell analyzer;

FIG. 12 is a flowchart illustrating a discrimination/substitutionprocessing;

FIG. 13 is a block diagram illustrating the internal configuration of acell analyzer according to the second embodiment;

FIG. 14 shows the first half of a flowchart illustrating the processingsperformed by the respective control sections of the cell analyzeraccording to the second embodiment; and

FIG. 15 shows the second half of a flowchart illustrating theprocessings performed by the respective control sections of the cellanalyzer according to the second embodiment.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

The following section will describe an embodiment of the cell analyzerand the cell analysis method of the present invention with reference tothe attached drawings.

[First Embodiment]

[Entire Configuration of Cell Analyzer]

FIG. 1 is a perspective view illustrating a cell analyzer 1 according tothe first embodiment of the present invention.

This cell analyzer 1 causes a measurement sample including cellsextracted from a patient to flow in a flow cell, emits laser beam to themeasurement sample flowing in the flow cell, detects the light from themeasurement sample (e.g., forward scattered light, side fluorescence) toanalyze the optical signal to thereby determine whether the cellsinclude a cancer cell or not.

More specifically, the cell analyzer 1 of the present embodiment is usedto analyze epidermal cells of a cervix and is used to screen a cervicalcancer.

As shown in FIG. 1, the cell analyzer 1 comprises: a measurementapparatus 2 for subjecting a measurement sample to an opticalmeasurement by laser beam; a sample preparation apparatus 3 forsubjecting a biological sample extracted from a subject to apreprocessing (e.g., cleaning or staining) to prepare a measurementsample to be supplied to the measurement apparatus 2; and a dataprocessing apparatus 4 for analyzing the measurement result by themeasurement apparatus 2 for example.

[Internal Configuration of Measurement Apparatus]

FIG. 2 is a block diagram illustrating the internal configuration of themeasurement apparatus 2.

As shown in FIG. 2, this measurement apparatus 2 comprises: a maindetecting section 6; a signal processing section 7; a measurementcontrol section 8; and an I/O interface 9.

Among these components, the main detecting section 6 detects, from ameasurement sample, the number and size of a measurement target cell aswell as the nucleus thereof for example. In the present embodiment, aflow cytometer 10 shown in FIG. 5 and FIG. 6 is used.

The signal processing section 7 is composed of a signal processingcircuit that performs a signal processing required for an output signalfrom the main detection section 6. The measurement control section 8includes a microprocessor 11 and a memory section 12. The memory section12 is composed of ROM and RAM for example.

The memory section 12 includes a ROM that stores therein controlprograms for controlling the operations of the main detecting section 6and the signal processing section 7 and data required to execute thecontrol programs. The microprocessor 11 can load the control programs toa RAM or can execute the programs directly from the ROM.

The microprocessor 11 of the measurement control section 8 is connected,via the I/O interface 9, to the data processing apparatus 4 and amicroprocessor 19 of a preparation control section 16 which will bedescribed later. Thus, the data processed by the microprocessor 11 andthe data required for the processing by the microprocessor 11 can besent and received between the data processing apparatus 4 and themicroprocessor 19 of the preparation control section 16.

[Internal Configuration of Sample Preparation Apparatus]

FIG. 3 is a block diagram illustrating the internal configuration of thesample preparation apparatus 3.

As shown in FIG. 3, this sample preparation apparatus 3 comprises: a subdetecting section 14; a signal processing section 15; the preparationcontrol section 16; an I/O interface 17; and a preparation devicesection 18 for automatically subjecting a biological sample to acomponent adjustment.

Among them, the sub detecting section 14 detects the number ofmeasurement target cells included in the biological sample. In thepresent embodiment, the sub detecting section 14 also uses substantiallythe same flow cytometer 10 as those shown in FIG. 5 and FIG. 6.

The signal processing section 15 is composed of a signal processingcircuit to perform a signal processing required for an output signalfrom the sub detecting section 14. The preparation control section 16 iscomposed of: the microprocessor 19; a memory section 20; a sensor driver21; and a driving section driver 22. The memory section 20 is composedof ROM and RAM for example.

The preparation device section 18 of the present embodiment is composedof: a specimen setting section 24; a cell dispersion section 25; aspecimen pipette section 26; a specimen quantitation section 27; areagent quantitation section 28; and a discrimination/substitutionsection 29.

Among them, the specimen setting section 24 is used to set a pluralityof living body containers 53 and product containers 54 (see FIG. 7) thatstore biological samples extracted from a patient and preservativesolution containing methanol as a major component. The cell dispersionsection 25 agitates mixed liquid of the biological sample and thepreservative solution in the living body container 53 to forcedlydisperse the cells included in the sample.

The specimen pipette section 26 is used to remove the mixed liquid ofthe biological sample and the preservative solution in which the cellsare dispersed from the living body container 53 to introduce the liquidto the fluid circuit of the preparation device section 18 or is used toreturn a prepared product to the product container 54 or to remove theproduct from the product container 54. The specimen quantitation section27 quantifies the mixed liquid of the biological sample and thepreservative solution supplied to the fluid circuit.

The reagent quantitation section 28 quantifies the reagent added to abiological sample such as staining fluid. Thediscrimination/substitution section 29 is used to mix the biologicalsample with the preservative solution and the diluted solution tosubstitute the preservative solution and the diluted solution, and todiscriminate the measurement target cell from cells other than themeasurement target cell (e.g., red blood cells, white blood cells) andbacteria for example. It is noted that the configuration of the fluidcircuit of the preparation device section 18 having the above respectivesections 24 to 29 (FIG. 7) will be described later.

The ROM of the memory section 20 stores therein the control programsused to control the operations of the sub detecting section 14, thesignal processing section 15, the sensor driver 21, and the drivingsection driver 22 and the data required to execute the control programs.The control program can be loaded by the microprocessor 19 to the RAMfor execution or can be directly executed from the ROM.

The microprocessor 19 of the preparation control section 16 isconnected, via the I/O interface 17, to the microprocessor 11 of themeasurement control section 8. Thus, the data processed by themicroprocessor 19 and the data required for the processing by themicroprocessor 19 can be sent and received between the microprocessor 19and the microprocessor 11 of the measurement processing section 8.

The microprocessor 19 of the preparation control section 16 isconnected, via the sensor driver 21 and the driving section driver 22,to the sensors or the like of the respective sections 24 to 29 of thepreparation device section 18 and a driving motor configuring a drivingsection. Based on a sensing signal from a sensor, the microprocessor 19executes a control program to control the operation of the drivingsection.

[Internal Configuration of Data Processing Section]

FIG. 4 is a block diagram illustrating the internal configuration of thedata processing apparatus 4.

As shown in FIG. 4, the data processing apparatus 4 of the presentembodiment is composed of, for example a personal computer such as alaptop PC (or a desk top PC) and is mainly composed of a processing body31, a display section 32, and an input section 33.

The processing body 31 comprises: a CPU 34; a ROM 35; a RAM 36; a harddisk 37; a reading apparatus 38; an input/output interface 39; and animage output interface 40. The respective sections are connected via aninternal bus so that these sections can communicate to one another.

The CPU 34 can execute a computer program memorized in the ROM 35 and acomputer program loaded to the RAM 36.

The ROM 35 is configured by a mask ROM, PROM, EPROM, EEPROM or the likeand stores therein a computer program executed by the CPU 34 and thedata used for the computer program for example.

The RAM 36 is configured by SRAM or DRAM for example. The RAM 36 is usedto read various computer programs recorded in the ROM 35 and the harddisk 37 and is used as a working region of the CPU 34 to execute thesecomputer programs.

In the hard disk 37, various computer programs to be executed by the CPU34 such as an operating system and an application program and the dataused to execute the programs are installed

In the hard disk 37, there is installed an operating system thatprovides, for example, a graphical user interface environment such asWindows® manufactured and sold by U.S. Microsoft Corporation.

Furthermore, in the hard disk 37, an operation program 41 is installedfor the transmission of an operation instruction to the measurementcontrol section 8 and the preparation control section 16, for thereception and analysis processings of the measurement result performedby the measurement apparatus 2, and for the display of the processedanalysis result. The operation program 41 operates on the operatingsystem.

The reading apparatus 38 is configured by a flexible disk drive, aCD-ROM drive, or a DVD-ROM drive and can read a computer program or datarecorded in a portable recording medium.

The input/output interface 39 is composed of, for example, a serialinterface such as USB, IEEE1394 or RS-232C, a parallel interface such asSCSI, IDE or IEEE1284, and an analog interface such as a D/A converteror an A/D converter.

The input/output interface 39 is connected to the input section 33composed of a keyboard and a mouse. By allowing a user to operate theinput section 33, data can be inputted to the computer.

The input/output interface 39 is also connected to the I/O interface 9of the measurement apparatus 2 to thereby provide data transmission andreception between the measurement apparatus 2 and the data processingapparatus 4.

The image output interface 40 is connected to the display section 32composed of LCD or CRT. The image output interface 40 causes the displaysection 32 to output a video signal depending on the image data from theCPU 34.

[Configuration of Main Detecting Section (Flow Cytometer)]

FIG. 5 is a function block diagram of the flow cytometer 10 configuringthe main detecting section 6. FIG. 6 is a side view illustrating theoptical system of the flow cytometer 10.

As shown in FIG. 5, the flow cytometer 10 has a lens system 43 thatcollects the laser beam from a semiconductor laser 44 as a light sourceto a measurement sample flowing in a flow cell 45. A collecting lens 46collects the forward scattered light from cells in the measurementsample to a scattered light detector composed of a photodiode 47.

Although FIG. 5 shows the lens system 43 by a single lens, the lenssystem 43 actually has a configuration as shown in FIG. 6 for example.

In other words, the lens system 43 of the present embodiment is composedof, in an order from the semiconductor laser 44 (left side of FIG. 6), acollimator lens 43 a, a cylinder lens system (a planoconvex cylinderlens 43 b+a biconcave cylinder lens 43 c) and a condenser lens system (acondenser lens 43 d+a condenser lens 43 e).

Returning to FIG. 5, a side collecting lens 48 collects the sidescattered light and the side fluorescence in the measurement target cellor the nucleus in the cell to a dichroic mirror 49. The mirror 49reflects the side scattered light to the photomultiplier 50 as ascattered light detector and transmits the side fluorescence to aphotomultiplier 51 as a fluorescence detector. These lights reflect thefeatures of the cell in the measurement sample or the nucleus.

Then, the photodiode 47 as well as the respective photomultipliers 50and 51 convert the received optical signal to an electric signal tooutput a forward scattered light signal (FSC), a side scattered lightsignal (SSC), and a side fluorescence signal (SFL), respectively. Theseoutput signals are amplified by a preamplifier (not shown) and is sentto the signal processing section 7 of the measurement apparatus 2 (FIG.2).

The respective signals FSC, SSC, and SFL processed by the signalprocessing section 7 of the measurement apparatus 2 are sent by themicroprocessor 11 from the I/O interface 9 to the data processingapparatus 4.

The CPU 34 of the data processing apparatus 4 executes the operationprogram 41 to thereby prepare a scattergram for analyzing the cell andnucleus based on the respective signals FSC, SSC, and SFL. Based on thescattergram, it is determined whether the cell in the measurement sampleis abnormal or not, specifically, whether the cell is a cancerous cellor not.

It is noted that, although the flow cytometer 10 may have a light sourcecomposed of a gas laser instead of the semiconductor laser 44, the useof the semiconductor laser 44 is preferred from the viewpoints of lowcost, small size, and low power consumption. The use of thesemiconductor laser 44 as described above can reduce the product costand can allow the apparatus to have a small size and power saving.

Furthermore, in the present embodiment, blue semiconductor laser havinga short wavelength is used that is advantageously used to narrow beam.The blue semiconductor laser is also advantageous to a fluorescenceexcitation wavelength such as PI. It is noted that red semiconductorlaser also may be used that is low-cost and long-life amongsemiconductor lasers and that can be supplied stably from manufacturers.

By the way, an epidermal cell of a cervix has an average size of about60 μm and the nucleus has a size of 5 to 7 μm. When the cell becomescancerous, the cell division frequency increases abnormally and thenucleus grows to a size of 10 to 15 μm. As a result, an N/C ratio(nucleus size/cell size) increases to a value larger than that of anormal cell.

Thus, by detecting the sizes of a cell and the nucleus, there can beobtained an indicator that is used to determine whether the cell iscancerous or not.

Thus, in the present embodiment, the scattered light from a measurementsample flowing in the flow cell 45 is detected by the photodiode 47 andthe fluorescence from the measurement sample flowing in the flow cell 45is detected by the photomultiplier 51.

The signal processing section 7 of the measurement apparatus 2 acquires,from the scattered light signal outputted from the photodiode 47, apulse width of the scattered light signal having a value reflecting thesize of the measurement target cell and acquires, from the fluorescencesignal outputted from the photomultiplier 51, a pulse width of thefluorescence signal having a value reflecting the size of the nucleus ofthe measurement target cell.

Then, based on the values obtained by the signal processing section 7that reflect the size of the measurement target cell and the size of thenucleus of the measurement target cell, whether the measurement targetcell is abnormal or not is determined by the CPU 34 of the dataprocessing apparatus 4 configuring an analysis section.

Specifically, the CPU 34 of the data processing apparatus 4 determinesthat the measurement target cell is abnormal when a value obtained bydividing the pulse width of the fluorescence signal by the pulse widthof the scattered light signal is larger than a predetermined thresholdvalue.

[Configuration of Sub Detecting Section]

The sub detection section 14 for performing preliminary detection of thesample preparation apparatus 3 detects, in a preprocessing step forpreparing a sample, the number of measurement target cells in abiological sample. In the present embodiment, the flow cytometer 10 isused that has substantially the same configuration as those of the flowcytometers illustratively shown in FIG. 5 and FIG. 6.

The sample preparation apparatus 3 of the present embodimentpreliminarily measures the concentration of measurement target cellsprior to the main measurement by the measurement apparatus 2.Accordingly, it is sufficient for the sub detecting section 14 to outputa signal for counting the number of the cells.

Thus, in the case of the flow cytometer 10 configuring the sub detectingsection 14, it is only required to acquire a forward scattered lightsignal (FSC). Thus, the photomultipliers 50 and 51 for acquiring a sidescattered light signal (SSC) and a side fluorescence signal (SFL) arenot provided and only the photodiode 47 for acquiring FSC is provided.

Then, the optical signal received by the photodiode 47 of the subdetecting section 14 as described above is converted to an electricsignal and is amplified. Then, the resultant signal is sent to thesignal processing section 15 of the sample preparation apparatus 3 (FIG.3).

The signal FSC processed by the signal processing section 15 of thesample preparation apparatus 3 is sent to the preparation controlsection 16. The microprocessor 19 of the preparation control section 16counts the number of the measurement target cells based on the signalFSC.

The microprocessor 19 acquires the volume of biological samplepreliminarily extracted by the specimen pipette section 26 forconcentration measurement from a flow rate sensor (not shown) providedin the pipette section 26. By dividing the number of the cells obtainedfrom the signal FSC by the volume, the concentration of the biologicalsample is calculated.

However, it is not always required to calculate the concentration itselfof the biological sample. When the extraction amount of the biologicalsample is retained at a fixed level, the number of cells itselffunctions as information reflecting the concentration thereof. In otherwords, the concentration information generated by the microprocessor 19of the preparation control section 16 may be not only the concentrationof the biological sample but also may be other information substantiallyequivalent to the concentration.

Then, the microprocessor 19 of the preparation control section 16issues, based on the above concentration information generated by themicroprocessor 19, control instructions for controlling the preparationoperation performed by the preparation device section 18 to thebiological sample (e.g., operation for quantifying the biological sampleor reagent). It is noted that specific contents of the control will bedescribed later.

[Fluid Circuit of Preparation Device Section]

FIG. 7 is a fluid circuit diagram of the preparation device section 18.

As shown in FIG. 7, the specimen setting section 24 comprises: acircular rotation table 24A; and a driving section 24B for driving therotation of the rotation table 24A. At the outer periphery of therotation table 24A, there is provided a retention section that can setthe living body container 53 for storing mixed liquid of the biologicalsample and preservative solution and a product container (microtube) 54for storing the product generated through thediscrimination/substitution processing by thediscrimination/substitution section 29.

The cell dispersion section 25 comprises: an agitation bar 25A foragitating the sample in the living body container 53; and a drivingsection 25B for driving the rotation of the agitation bar 25A. Thedriving section 25B inserts the agitation bar 25A into the living bodycontainer 53 to rotate the agitation bar 25A.

As a result, the biological sample in the living body container 53 isagitated and the cells included in the biological sample can bedispersed.

The specimen pipette section 26 comprises: the first pipette 26A thatsucks the biological sample in the living body container 53 to supplythe sample to the specimen quantitation section 27 and to return theproduct generated by the discrimination/substitution section 29 to theproduct container 54; and the second pipette 26B that supplies reagentsuch as staining fluid quantified by the reagent quantitation section 28to the product container 54.

The first pipette 26A is connected via a pipe line to a storagecontainer 57 of the discrimination/substitution section 29 which will bedescribed later. Thus, liquid for which the measurement target cells arediscriminated by the discrimination/substitution section 29 can bereturned by the first pipette 26A to the product container 54.

The specimen quantitation section 27 comprises: a quantitation cylinder27A; and a driving section 27B for moving a quantitation piston insertedin the cylinder 27A in the up-and-down direction. The quantitationcylinder 27A is connected by a pipe line to the first pipette 26A via adirection switching valve V1.

The mixed liquid of the biological sample and the preservative solutionsucked from the living body container 53 via the first pipette 26A isintroduced to the quantitation cylinder 27A through the directionswitching valve V1. The introduced mixed liquid of the biological sampleand the preservative solution is sent to the nextdiscrimination/substitution section 29 through the direction switchingvalve V1 by the travel of the quantitation piston by the driving section27B.

It is noted that, in the present embodiment, the quantitation cylinder27A of the specimen quantitation section 27 is also connected via a pipeline to a diluted solution unit 55 for preparing diluted solution for abiological sample.

The reagent quantitation section 28 comprises: a pair of quantitationcylinders 28A and 28B; and a driving section 28C for moving quantitationpitons respectively inserted to the cylinders 28A and 28B in theup-and-down direction. The cylinders 28A and 28B are connected by a pipeline to the second pipette 26B via supply switching valves V2 and V3,respectively.

The reagent in the reagent container is supplied to the respectivequantitation cylinders 28A and 28B. The supplied reagent is quantifiedin a predetermined amount by the travel of the quantitation piston bythe driving section 28C. Then, the quantified reagent is sent to thesecond pipette 26B via the supply switching valves V2 and V3.

Thus, the discriminated sample returned to the product container 54 ofthe specimen setting section 24 can be mixed with a plurality of typesof reagents in predetermined amounts quantified by the reagentquantitation section 28.

In the present embodiment, there are two types of reagents quantified bythe respective quantitation cylinders 28A and 28B of the reagentquantitation section 28. Among these reagents, the reagent that ismeasured by one quantitation cylinder 28A and that is added to abiological sample is staining liquid for performing PI staining. Thereagent that is measured by the other quantitation cylinder 28B and thatis added to the biological sample is RNase for subjecting cells to anRNA processing. The PI staining is performed by propidium iodide (PI)that is fluorescence staining fluid including dye. In the PI staining, anucleus is selectively stained. Thus, the fluorescence from the nucleuscan be detected. The RNA processing is a processing for dissolving RNAin the cell. The staining liquid stains both of the RNA and DNA of anepidermal cell. Thus, RNA is dissolved by performing the above RNAprocessing and is not stained by the staining liquid, so that the cellnucleus DNA can be measured accurately.

The discrimination/substitution section 29 comprises: the storagecontainer 57 having an opening shape at the upper part; a filtrationcylinder 58 inserted to the storage container 57 so as to be movable inthe up-and-down direction; and a driving section 59 for moving thefiltration cylinder 58 in the up-and-down direction in the storagecontainer 57.

The storage container 57 is connected by a pipe line to the quantitationcylinder 27A of the specimen quantitation section 27 via the directionswitching valves V1, V7, and V4. Thus, the mixed liquid of thebiological sample and the preservative solution quantified by thespecimen quantitation section 27 is supplied via the direction switchingvalves V1, V7, and V4 to the storage container 57 and can be onceretained in the interior of the container 57. The already-discriminatedbiological sample in the storage container 57 is sent via the same pathto the first pipette 26A.

Furthermore, the storage container 57 is connected to the exterior ofthe storage container 57 via the switching valve V9. By opening theswitching valve V9, the interior of the storage container 57 can beopened to air.

The pipe line part from the direction switching valve V4 to the disposalsection 61 includes the flow cell 45 of the sub detecting section 14.Thus, the sub detecting section 14 can count the number of the cells ofthe already-discriminated biological sample discharged from the storagecontainer 57.

The filtration cylinder 58 is composed of a hollow tube having a filter60 at the lower part that does not allow a measurement target cell(epidermal cell) to pass therethrough and that allows cells having asmaller diameter than that of the measurement target cell (e.g., redblood cells, white blood cells) to pass therethrough. The filtrationcylinder 58 is connected via a pipe line to the diluted solution unit 55via the switching valve V5.

Thus, the diluted solution of the diluted solution unit 55 can besupplied to the interior of the filtration cylinder 58 by opening theswitching valve V5.

The driving section 59 of the discrimination/substitution section 29causes the filtration cylinder 58 to travel in the downward direction tothe storage container 57 containing therein the mixed liquid of thebiological sample, the preservative solution, and the diluted solution.Then, the filter 60 of the filtration cylinder 58 downwardly moves inthe mixed liquid in the storage container 57. As a result, liquid mainlyincluding the measurement target cell remains as residual liquid at thelower part of the filter 60. At the same time, liquid mainly includingthe other cells and foreign substances remains as filtrate at the upperpart of the filter 60 (the interior of the filtration cylinder 58).

The filtration cylinder 58 is connected by a pipe line via the switchingvalve V6 to the disposal section 61 of the filtrate. Thus, the filtratefiltrated by the lowering of the filtration cylinder 58 is disposed tothe outside through the switching valve V6.

On the other hand, when the residual liquid after the filtration is formeasuring the concentration of the biological sample, the residualliquid in the filtration cylinder 58 is sent, after the disposal of thefiltrate, via the direction switching valve V4 to the flow cell 45 ofthe sub detecting section 14. Thereafter, the residual liquid isdisposed to the disposal section 61. When the residual liquid is forpreparing a measurement sample, the residual liquid is returned from thefirst pipette 26A to the product container 54.

As shown in FIG. 7, the quantitation cylinder 27A of the specimenquantitation section 27 is also connected by a pipe line to the maindetecting section 6 of the measurement apparatus 2 via the directionswitching valves V1 and V7.

The measurement sample in the product container 54 is quantified by thespecimen quantitation section 27 via the first pipette 26A. Then, thesample is supplied via the valves V1 and V7 to the main detectingsection 6 of the measurement apparatus 2.

It is noted that the operations of the driving section of the respectivesections and the switching valves (magnet valves) V1 to V7 shown in FIG.7 are controlled based on a control instruction from the preparationcontrol section 16 (the microprocessor 19). The preprocessing stepperformed by the preparation control section 16 will be described later.

[Specific Configuration of Discrimination/Substitution Section]

FIG. 8 is an expanded cross-sectional view illustrating a more specificconfiguration of the discrimination/substitution section 29. FIG. 9 isan explanatory diagram illustrating the filtration function thereof.

As shown in FIG. 8, the discrimination/substitution section 29 of thepresent embodiment comprises: the storage container 57 that can containtherein liquid L including a biological sample; and the filter 60 thatdoes not allow the first cell C1 in the biological sample to passtherethrough and that allows the second cell C2 having a smallerdiameter than that of the first cell C1 to pass therethrough.

Furthermore, the discrimination/substitution section 29 comprises thefiltration cylinder 58 as a liquid separation section that allows theliquid L to pass the filter 60 to separate the liquid L into the firstliquid L1 mainly including the first cell C1 and the second liquid L2mainly including the second cell C2.

The storage container 57 has, in an integrated manner, a hollow bodysection 57A having a shaft center in the up-and-down direction and abottom section 57B that is connected to the lower part of the bodysection 57A and that has a flat inner surface of which center part isconcave to the lower direction.

Furthermore, the discrimination/substitution section 29 comprises, inthe vicinity of the inner face of the bottom section 57B of the storagecontainer 57, a bar-like stirrer bar (stirrer) 68 rotating by a magneticforce and, at the lower side of the bottom section 57B, a magnet 69 forgiving a magnetic force to the stirrer bar 68 and a driving motor 70 forrotating the magnet 69. Thus, by rotating the magnet 69 by the drivingmotor 70 to thereby rotate the stirrer bar 68, the liquid in the storagecontainer 57 can be agitated.

It is noted that the filtration cylinder 58 is connected to a positivepressure source 71 via the switching valve V8. Thus, by opening theswitching valve V8, a positive pressure can be supplied to the interiorof the filtration cylinder 58.

On the other hand, the filtration cylinder 58 comprises: a hollowtube-like cylinder 63 inserted into the storage container 57 via a sealmember 62 so as to be movable in the up-and-down direction; and thefilter 60 provided so as to seal the lower end opening of the cylinder63.

In the present embodiment, the first cell C1 is assumed as an epidermalcell of a cervix. This epidermal cell has a size of about 20 to 80 μm(an average size is about 60 μm). A red blood cell as the second cellC2, which is smaller than the first cell C1, has a size of about 7 to 10μm. A white blood cell as other second cell C2 has a size of about 8 to15 μm. Foreign substance such as bacteria as other second cell C2 has asize of 1 to a few μm.

Thus, in the present embodiment, in order to prevent an epidermal cellfrom passing the through hole of the filter 60 even when a pressure isapplied to the liquid in the storage container 57, the filter 60 is ametal CVD (Chemical Vapor Deposition) filter with a through hole havinga diameter of 8 to 20 μm smaller than 20 μm. In the case of the metalCVD filter as described above, the through hole thereof is suppressedfrom being deformed when compared with the other resin filters or metalmesh filters and the aperture ratio can be advantageously increased.

Furthermore, the reason why the hole diameter of the filter 60 is set to8 to 20 μm is that a diameter smaller than 8 μm frequently shows aphenomenon where the through hole is clogged with a cell or foreignsubstance at an early stage and a diameter exceeding 20 μm on the otherhand frequently causes an epidermal cell to undesirably pass through athrough hole when a pressure is applied to the liquid in the storagecontainer 57. It is noted that the hole of the filter 60 preferably hasa diameter of about 15μ.

The bottom section 57B of the storage container 57 is connected to aresidual liquid pipe line 64 for introducing the already quantifiedbiological sample and for discharging, to the outside, the residualliquid L1 for which filtration is already performed to acquire theresultant liquid. At a middle of this pipe line 64, the directionswitching valve V4 is provided. Thus, the residual liquid pipe line 64and the direction switching valve V4 configure a liquid acquisitionsection that is used to acquire the first liquid L1 separated by thefiltration cylinder 58 functioning as a liquid separation section.

The upper wall section of the cylinder 63 is connected to the dilutedsolution pipe line 65 leading to the diluted solution unit 55. At amiddle of this pipe line 65, the switching valve V5 is provided.

The upper wall section of the cylinder 63 is connected to the filtratepipe line 66 that is used to dispose the filtrated filtrate L2 to theoutside and that leads to the disposal section 61. At a middle of thepipe line 66, the switching valve V6 is provided. Thus, the filtratepipe line 66 and the switching valve V6 configure a liquid dischargesection for discharging, to the outside, the second liquid L2 separatedby the filtration cylinder 58 functioning as a liquid separationsection.

An upper end of the cylinder 63 is connected to a travel mechanismsection 67 that is used to convert the rotation movement of the drivingsection 59 composed of a motor for example to the travel in theup-and-down direction of the cylinder 63.

The driving section 59 of the travel mechanism section 67 drives thefiltration cylinder 58 in accordance with a control instruction from thepreparation control section 16 (microprocessor 19).

For example, as shown in FIG. 9(a), the preparation control section 16lowers the filtration cylinder 58 so that the filter 60 travels in thelower direction from the upper side of the liquid level of the liquid Lin the storage container 57 (the mixed liquid of the biological sample,the preservative solution, and the diluted solution) to the interior ofthe liquid.

Then, as shown in FIG. 9(b), the liquid in which almost all cellsincluded therein are the first cells (epidermal cell) C1 remains as theresidual liquid L1 at the lower part of the filter 60 in the storagecontainer 57. The liquid including the other second cells C2 (foreignsubstances such as red blood cells, white blood cells, and bacteria)remains as the filtrate L2 at the upper part of the filter 60 (theinterior of the filtration cylinder 58).

Thereafter, the preparation control section 16 controls the drivingsection 59 of the travel mechanism section 65 so that the filter 60already travelled into the liquid L is returned in the upper directionby a predetermined distance.

Specifically, the preparation control section 16 controls the travelmechanism section 67 so that the filtration cylinder 58 is lowered untilthe filter 60 reaches a lowering limit set at the bottom section 57B orin the vicinity thereof and then the filter 60 is returned in the upwarddirection by a predetermined distance. By returning in the upwarddirection, the first cell C1 attached to the lower face of the filter 60can be separated from the filter 60.

Then, the preparation control section 16 firstly releases the switchingvalve V6. As a result, as shown in FIG. 9(c), the filtrate L2 isdischarged to the outside earlier than the residual liquid L1.Thereafter, the direction switching valve V4 is released in order toacquire the residual liquid L1.

[Contents of Analysis Processing]

FIG. 10 and FIG. 11 are a flowchart illustrating the processingsperformed by the respective control sections 8, 16, and 31 of the cellanalyzer 1.

It is noted that FIG. 10 shows, at the right column, a processing flowperformed by the control section (processing body) 31 of the dataprocessing apparatus 4 and shows, at the left column, a processing flowperformed by the control section 8 of the measurement apparatus 2. FIG.11 shows a processing flow performed by the control section 16 of thesample preparation apparatus 3 in a single column. However, thisprocessing flow is connected to the processing flow of FIG. 10 at theshown points A, B, and C. The following section will describe, withreference to FIG. 10 and FIG. 11, the processing contents performed bythe cell analyzer 1.

First, the control section 31 of the data processing apparatus 4 causesthe display section 32 to display a menu screen (Step S1). Thereafter,upon accepting a measurement starting instruction based on the menuscreen from the input section 33 (Step S2), the control section 31 ofthe data processing apparatus 4 sends a measurement starting signal tothe measurement apparatus 2 (Step S3).

Upon receiving the measurement starting signal (Step S4), the controlsection 8 of the measurement apparatus 2 sends a preparation startingsignal to the sample preparation apparatus 3 (Step S5 and point A).

Upon receiving the preparation starting signal (Step S6), the controlsection 16 of the sample preparation apparatus 3 sucks the reagent(staining fluid, RNase) used to prepare a measurement sample into a flowpath in the apparatus and disperses, in the cell dispersion section 25,the cells in the mixed liquid of the biological sample and preservativesolution including methanol as a major component contained in the livingbody container 53 (Steps S7 and S8).

Thereafter, the control section 16 of the sample preparation apparatus 3causes the already-dispersed mixed liquid to be sucked by apredetermined amount from the living body container 53 into the flowpath in the apparatus (Step S9) and causes the liquid to be sent to thestorage container 57 of the discrimination/substitution section 29.Then, the discrimination/substitution section 29 is caused to perform adiscrimination/substitution processing to the mixed liquid of thebiological sample and the preservative solution (Step S10).

[Contents of Discrimination/Substitution Processing]

FIG. 12 is a flowchart illustrating the abovediscrimination/substitution processing.

As shown in FIG. 12, the control section 16 of the sample preparationapparatus 3 firstly causes diluted solution to be mixed with the mixedliquid of the biological sample and the preservative solution in thestorage container 57 (Step T1). More specifically, the mixed liquid issupplied to the storage container 57. Then, the diluted solution isinputted to the cylinder 63 at a position slightly upper side of thelower limit. Next, the cylinder 63 is raised to the upper limitposition. This consequently mixes the mixed liquid with the dilutedsolution.

Next, the cylinder 63 is lowered to the lower limit position (Step T2).By the lowering of the cylinder 63, the liquid L in the storagecontainer 57 is discriminated, as described above, to a predeterminedamount of the residual liquid L1 mainly including the first cell C1 as ameasurement target and the filtrate L2 mainly including the cells C2other than the first cell C1.

Thereafter, the control section 16 of the sample preparation apparatus 3causes, at a time point at which the cylinder 63 reaches the lowerlimit, the cylinder 63 to be raised by a predetermined distance (StepT3). As a result, the first cell (epidermal cell) C1 attached to thelower face of the filter 60 can be floated in the concentrated liquid L1at the lower side of the filter 60.

Thus, the travel in the processing (Step T3) for which the cylinder 63is raised may be performed at such speed and stroke that allows thefirst cell C1 to be physically separated from the filter 60. The travelmay be performed a plurality of times. After the above processing, thecontrol section 16 of the sample preparation apparatus 3 discharges thefiltrate L2 existing above the filter 60 to the outside (Step T4).

Next, the control section 16 of the sample preparation apparatus 3 usesthe driving motor 70 to rotate the magnet 69 to thereby rotate thestirrer bar 68 in the storage container 57 (Step T5). The rotation ofthe stirrer bar 68 causes a circular flow in the horizontal direction inthe concentrated liquid L1 in the storage container 57, thereby causing,in the lower face of the filter 60, a shearing force in the horizontaldirection. Thus, the first cell (epidermal cell) C1 attached to thelower face of the filter 60 can be removed by the above shearing forceand can be floated in the concentrated liquid L1 at the lower side ofthe filter 60. A distance between the lower face of the filter 60(filtering area) and the filter 60-side face of the stirrer bar 68opposed to the lower face is not particularly limited but is preferable1 mm or less and more preferably 0.6 mm or less. The rotation number ofthe magnet 69 for rotating the stirrer bar 68 (i.e., the rotation numberof the driving motor 70) is preferably in a range from 1000 to 2000 rpmand is more preferably about 1300 rpm.

Next, the control section 16 opens the switching valve V9 to open, toair, the space between the inner periphery face of the body section 57Aof the storage container 57 and the outer periphery face of the cylinder63 and opens the switching valve V6 to apply a negative pressure to theinterior of the filtration cylinder 58 (Step T6). During thisprocessing, the disposal section 61 is configured to function as anegative pressure source. This can consequently cause the concentratedliquid L1 existing between the inner periphery face of the body section57A of the storage container 57 and the outer periphery face of thecylinder 63 to travel to the lower side of the filter 60. As a result, asmall liquid part exists at the upper side of the filter 60.

Next, the control section 16 closes the switching valve V6 and thenopens the switching valve V8 to apply positive pressure (e.g., 15 kPa)from the positive pressure source 71 into the filtration cylinder 58 fora predetermined time (e.g., 0.1 second) (Step T7). As a result, apressure can be applied from the upper side of the filter 60 to thethrough hole of the filter 60. Thus, the first cell (epidermal cell) C1clogging the through hole of the filter 60 can be removed and can befloated in the concentrated liquid L1 at the lower side of the filter60. Furthermore, since a pressure is applied from the upper side of thefilter 60 while a small liquid part being existing at the upper side ofthe filter 60, a pressure can be uniformly applied to a plurality ofthrough holes of the filter 60 when compared with a case where apressure is directly applied to the filter 60 without passing through aliquid part. Thus, the clogging of the filter 60 can be suppressedeffectively.

Thereafter, the switching valves V8 and V9 are closed (Step T8). Then,the rotation of the stirrer bar 68 is stopped (Step T9).

Next, the control section 16 of the sample preparation apparatus 3determines whether the lowering of the cylinder 63 is the second one ornot (Step T10).

When the lowering of the cylinder 63 is not the second one, the controlsection 16 of the sample preparation apparatus 3 repeats the mixing ofthe diluted solution to the filtration. When the lowering of thecylinder 63 is the second one, the control section 16 of the samplepreparation apparatus 3 completes the routine of thediscrimination/substitution processing. It is noted that the number atwhich this discrimination/substitution operation is performed is notlimited to two and also may be a number larger than two.

By the discrimination/substitution processing, there can be acquiredsuch liquid that mainly includes the first cell (epidermal cell) C1 as ameasurement target cell and that includes a reduced number of the secondcells C2 other than the measurement target cell. Furthermore, by theabove discrimination/substitution processing, the concentration of thepreservative solution in the liquid (the mixed liquid of the biologicalsample and the preservative solution) supplied from the living bodycontainer 53 to the storage container 57 can be reduced by substitutingthe most part of the preservation solution with the diluted solution.Thus, in a DNA staining processing which will be described later, theinfluence by the preservative solution can be reduced and the DNA of themeasurement target cell can be stained favorably.

Furthermore, in the discrimination/substitution processing, thesubstitution processing of the preservative solution and the dilutedsolution can be performed while the cell discrimination processing isbeing performed. Thus, the discrimination processing and thesubstitution processing can be performed with a shorter time whencompared with a case where these two processings are performedseparately.

Furthermore, in the discrimination/substitution processing, the stirrerbar 68 is rotated to remove the first cell (epidermal cell) C1 attachedto the lower face of the filter 60 by a shearing force to float thefirst cell (epidermal cell) C1 in the concentrated liquid L1 at thelower side of the filter 60. A pressure is applied from the upper sideof the filter 60 to the through hole of the filter 60 to thereby removethe first cell (epidermal cell) C1 clogging the through hole of thefilter 60, thus allowing the first cell (epidermal cell) C1 to float inthe concentrated liquid L1 at the lower side of the filter 60. Thus, thefirst cell (epidermal cell) C1 attached to the filter can be efficientlycollected without loss.

Table 1 shows data showing to which level the epidermal cell collectionrate is improved when the rotation of the stirrer bar and/or theapplication of a positive pressure to the through hole of the filterare/is performed.

The application of a positive pressure was performed by applying apressure of 15 kPa for 0.1 second. Such a stirrer bar was used that iscomposed of a circular disk in which an upper face includes across-shaped convex section. This stirrer bar was rotated at 1400 rpm.

TABLE 1 Rotation of stirrer Application of positive Number of singleIncrease rate of single Sample type bar [1400 rpm] pressure [15 kPa]epidermal cells epidermal cells [%] Example 1 A Yes No 37136 210 Example2 A Yes Yes 40950 231 Example 3 A No Yes 23890 135 Example 4 B Yes No55668 143 Example 5 B Yes Yes 63810 164 Example 6 B No Yes 52596 135Comparative Example 1 A No No 17692 100 Comparative Example 1 B No No38996 100

As shown in Example 6 of Table 1, the application of a positive pressureof 15 kPa provides an increase of the epidermal cell collection rate ofabout 35% when compared with a case where both of the positive pressureand the stirrer bar rotation are not used. Furthermore, the rotation ofthe stirrer bar at 1400 rpm (Examples 1, 2, 4, and 5) provides anincrease of the epidermal cell collection rate of about 43 to 110% whencompared with a case where both of the positive pressure and the stirrerbar rotation are not used.

Returning to FIG. 11, when the above discrimination/substitutionprocessing (Step S10) is completed, the control section 16 of the samplepreparation apparatus 3 sends to the flow cell 45 of the sub detectingsection 14 the concentrated liquid (the residual liquid L1 in which mostof cells included therein are epidermal cells) (Step S11). Then, thecontrol section 16 uses the sub detecting section 14 to subject theconcentrated liquid L1 to a premeasurement by a flow cytometry technique(Step S12).

This premeasurement is to obtain, prior to the main measurementperformed by the measurement apparatus 2 for cancer determination, suchconcentration information that reflects the concentration of themeasurement target cells (epidermal cells) C1 included in the biologicalsample. For example, this premeasurement is performed by detecting thenumber of the cells C1 included in the concentrated liquid L1.

[Use of Concentration Information]

Next, the control section 16 of the sample preparation apparatus 3discharges the concentrated liquid L1 to the disposal section 61 (StepS13) and calculates the concentration of the biological sample (StepS14). Then, based on the concentration information, the control section16 determines the suction amount of the biological sample for preparinga measurement sample for the main measurement (Step S15).

For example, when it is assumed that the suction amount of thebiological sample used for the premeasurement is 200 μl and thepremeasurement result shows that the number of the cells C1 is 10,000,the concentration of biological sample can be calculated as 10000/200μl=50 (number of cells/μl)=50000 (number of cells/ml).

On the other hand, when assuming that the number of significant cellsrequired to detect cancer cells in the main measurement is 100,000 forexample, the extraction amount of the biological sample required toperform the main measurement so as to secure the number of thesignificant cells may be computed as 100,000÷50000 (number ofcells/ml)=2 ml.

Thus, the control section 16 of the sample preparation apparatus 3 sucksthe biological sample from the living body container 53 by the requiredamount (Step S16). Then, the control section 16 subjects the biologicalsample to the above-described discrimination/substitution processingagain (Step S17).

As described above, the control section 16 of the sample preparationapparatus 3 determines, based on the concentration information for thecell C1 generated by the control section 16, the amount of thebiological sample for the preparation of the measurement sample used forthe main measurement and controls the preparation device section 18 ofthe sample preparation apparatus 3 so as to acquire the determinedamount of the biological sample.

In this case, the control section 16 of the sample preparation apparatus3 determines a higher amount of the biological sample used for thepreparation of the measurement sample as the cells C1 in the biologicalsample has a lower concentration and determines a lower amount of thebiological sample used for the preparation of the measurement sample asthe cells C1 in the biological sample has a higher concentration.

It is noted that, in the control section 16 of the sample preparationapparatus 3, the number of the cells C1 detected by the main detectingsection 34 is not limited to the above 100,000 as the number ofsignificant cells for cancer detection. Thus, the amount of thebiological sample for the preparation of the measurement sample may bedetermined so as to be within a predetermined range such as 100,000 to150,000.

Furthermore, the control section 16 of the sample preparation apparatus3 also can determine, based on the concentration information for thecells C1 generated by the control section 16, not only the amount of thebiological sample but also the amount of reagent for the preparation ofthe measurement sample. Then, the control section 16 can control thepreparation device section 18 of the sample preparation apparatus 3 soas to acquire the determined amount of reagent.

Furthermore, the control section 16 of the sample preparation apparatus3 also can determine, based on the concentration information for thecells C1 generated by the control section 16, the amount of at least oneof the biological sample and the reagent so that the ratio between thenumber of the cells C1 in the biological sample and the amount of thereagent (e.g., staining fluid) is within a predetermined range. Then,the control section 16 can control the preparation device section 18 ofthe sample preparation apparatus 3 so as to acquire the determinedamount of the biological sample or the reagent.

[Preparation of Measurement Sample]

Next, the control section 16 of the sample preparation apparatus 3supplies, to a product container (microtube) 54, the concentrated liquidL1 obtained through the second discrimination/substitution processing(Step S18) and supplies the staining fluid and RNase stored in theapparatus from the reagent quantitation section 28 to the productcontainer 54 (Step S19). Then, the control section 16 causes DNAstaining and RNA processing to be performed in the product container 54to thereby prepare a measurement sample (Step S20).

After the above processing, the control section 16 of the samplepreparation apparatus 3 sends the resultant measurement sample to themain detecting section 6 of the measurement apparatus 2 (Step S21 andpoint B).

It is noted that the control section 16 of the sample preparationapparatus 3 always determines whether a shutdown signal from themeasurement apparatus 2 is received or not (Step S22 and point C). Whenthe signal is not received, the processing returns to Step S6 fordetermining whether the preparation starting signal is received or not.When the signal is received, the processing executes the shutdown tothereby complete the sample preparation processing (Step S23).

[Main Measurement by Measurement Apparatus and Analysis of ResultantData]

Returning to FIG. 10, the control section 8 of the measurement apparatus2 always determines, after sending a preparation starting signal,whether the measurement sample is supplied from the sample preparationapparatus 3 or not (Step S24).

Thus, when the measurement sample is sent from the sample preparationapparatus 3 (point B), the control section 8 of the measurementapparatus 2 sends the measurement sample to the flow cell 45 of the mainmeasurement section 14 and subjects the cell C1 of the measurementsample to the main measurement (Step S25). Then, the control section 8sends the measurement data to the data processing apparatus 4 (StepS26).

On the other hand, the control section 31 of the data processingapparatus 4 always determines, after sending the measurement startingsignal, whether the measurement data is received from the measurementapparatus 2 or not (Step S27).

Upon receiving the above measurement data from the measurement apparatus2, the control section 31 of the data processing apparatus 4 uses themeasurement data to analyze the cell or nucleus to determine whether thecell in the measurement sample is cancerous or not for example (StepS28).

The control section 31 of the data processing apparatus 4 causes theabove analysis result to be displayed on the display section 32 (StepS29) and determines whether there is a shutdown instruction by userinput or not (Step S30).

When there is the above shutdown instruction, the control section 31 ofthe data processing apparatus 4 sends a shutdown signal to themeasurement apparatus 2 (Step S31).

The control section 8 of the measurement apparatus 2 always determineswhether the above shutdown signal from the data processing apparatus 4is received or not (Step S32). When the signal is not received, theprocessing returns to Step S4 for determining whether a measurementstarting signal is received or not. When the signal is received, theabove shutdown signal is transferred to the sample preparation apparatus3 (Step S33) and the shutdown is executed to thereby complete themeasurement processing (Step S34).

As described above, according to the cell analyzer 1 of the presentembodiment, liquid mainly including the first cells C1 having a largerdiameter is directly acquired as liquid. This can consequently eliminatethe need to separate measurement target cells captured by a filter fromthe filter to collect the cells, thus easily collecting the first cellsC1 as measurement target cells.

Thus, measurement target cells discriminated by a filter from othercells can be easily collected. At the same time, the use of themeasurement target cells thus collected can provide an easy preparationof a measurement sample suitable for the analysis of a measurementtarget cell by the sample preparation apparatus 3. Thus, the measurementresult also can be acquired for which an influence by cells other thanthe measurement target cell is reduced and the measurement target cellcan be analyzed accurately.

[Second Embodiment]

FIG. 13 is a block diagram illustrating the internal configuration ofthe cell analyzer 1 according to the second embodiment of the presentinvention.

The cell analyzer 1 of the second embodiment is different from that ofthe first embodiment in that all devices required for the premeasurementperformed by the sample preparation apparatus 3 are integratedlyprovided in the interior of the measurement apparatus 2. The respectivecomponents have the same configurations and functions as those in thefirst embodiment.

Thus, the configurations and functions of the respective sections of themeasurement apparatus 2 will not be described further by attaching thereference numerals used in the first embodiment also to the respectivesections of FIG. 13.

FIG. 14 and FIG. 15 are the first half and the second half of aflowchart illustrating the processings performed by the respectivecontrol sections 8 and 31 of the cell analyzer 1 according to the secondembodiment, respectively. It is noted that D to I of FIG. 14 areconnected to D to I of FIG. 15, respectively.

As shown in FIG. 14, the control section 31 of the data processingapparatus 4 firstly causes the display section 32 to display a menuscreen (Step S1). Thereafter, when a measurement starting instructionbased on the menu screen is accepted from the input section 33 (StepS2), the control section 31 of the data processing apparatus 4 sends ameasurement starting signal to the measurement apparatus 2 (Step S3).

Then, in the present embodiment, the control section 8 of themeasurement apparatus 2 also performs the processing performed in thefirst embodiment by the control section 16 of the sample preparationapparatus 3.

In other words, when the control section 8 of the measurement apparatus2 receives a measurement starting signal (Step S4), the control section8 sucks the reagent used for the preparation of a measurement sample(staining fluid, RNase) into the flow path in the apparatus. The controlsection 8 also causes, in the cell dispersion section 25, the dispersionof the cells in mixed liquid of a biological sample and preservativesolution including methanol as a major component contained in the livingbody container 53 (Steps S7 and S8).

Thereafter, the control section 8 of the measurement apparatus 2performs the same processing as in the control section 16 of the samplepreparation apparatus 3 in the first embodiment until the measurementsample is sent to the main detecting section 6 (Steps S7 to S21).

Then, the control section 8 of the measurement apparatus 2 performs,after sending the measurement sample to the flow cell 45 of the maindetecting section 6, the main measurement to the cell C1 of themeasurement sample (Step S25) and sends the measurement data to the dataprocessing apparatus 4 (Step S26).

On the other hand, the control section 31 of the data processingapparatus 4 always determines, after sending the measurement startingsignal, whether the measurement data is received from the measurementapparatus 2 or not (Step S27).

Upon receiving the above measurement data from the measurement apparatus2, the control section 31 of the data processing apparatus 4 uses themeasurement data to analyze the cell or nucleus to determine whether thecell in the measurement sample is cancerous or not for example (StepS28).

Furthermore, the control section 31 of the data processing apparatus 4causes the above analysis result to be displayed on the display section32 (Step S29) and determines whether there is a shutdown instruction byuser input or not (Step S30).

When there is the above shutdown instruction, the control section 31 ofthe data processing apparatus 4 sends a shutdown signal to themeasurement apparatus 2 (Step S31).

The control section 8 of the measurement apparatus 2 always determineswhether the above shutdown signal from the data processing apparatus 4is received or not (Step S32). When the signal is not received, then theprocessing returns to Step S4 for determining whether a measurementstarting signal is received or not. When the signal is received, theshutdown is executed to thereby complete the measurement processing(Step S34).

[Other Modified Examples]

It is noted that the above disclosed embodiment is an illustration ofthe present invention and is not the limited one. The scope of thepresent invention is shown not by the above embodiment but by the claimsand includes all modifications equivalent to the configurations in theclaims.

For example, the reagent storage section is not required to be includedin the fluid circuit in the apparatus as shown in FIG. 7. Thus, anotherconfiguration also may be used where reagent in a storage sectionexterior to the apparatus is introduced via the reagent quantitationsection 28 to the fluid circuit.

Furthermore, in the above embodiment, after the concentration of theepidermal cell C1 of a cervix is measured, a biological sample is newlysucked through the living body container 53 to supply the sample to thediscrimination/substitution section 29, and the concentrated liquid C1discriminated again is used to prepare a measurement sample. However,the concentrated liquid C1 used for the concentration measurement of theepidermal cell C1 also can be directly used as material of a measurementsample.

Furthermore, although the epidermal cell C1 of a cervix is assumed as ameasurement target cell in the above embodiment, cancerous determinationalso can be performed on buccal cells, other epidermal cells such asbladder and pharyngeals as well as epidermal cells of organs.

Furthermore, in the above embodiment, the main detecting section 6 andthe sub detecting section 14 are provided separately. However, anapparatus configuration also can be used where these detecting sections6 and 14 are both provided by one flow cytometer 10.

Furthermore, the sub detecting section 14 for counting the number of thecell C1 in the biological sample is not limited to the optical one suchas the flow cytometer 10 and also may be an electric resistance-typecell detector.

Furthermore, in the above embodiment, the liquid in the living bodycontainer 53 (the mixed liquid of the biological sample and thepreservative solution) is sucked by the pipette 26A and the suckedliquid is supplied via a pipe line to the storage container 57 of thediscrimination/substitution section 29. However, another configurationalso may be used where the pipette 26A is configured as the mobile one,and the liquid in the living body container 53 is sucked by the pipette26A and then the pipette 26A is caused to travel to the storagecontainer 57 of the discrimination/substitution section 29 tosubsequently discharge the liquid from the pipette 26A to the storagecontainer 57.

Furthermore, the discrimination/substitution section 29 uses a closedsystem and the filtration cylinder 58 is allowed to travel in thestorage container 57 in the up-and-down direction to thereby separatethe liquid L in the storage container 57 into the first liquid L1 andthe second liquid L2. However, another configuration also may be usedwhere the discrimination/substitution section 29 uses an open system anda negative pressure is applied to the interior of the filtrationcylinder 58 and the filtration cylinder 58 is allowed to travel in thedownward direction so as to follow the rise of the liquid level of thefirst liquid L1, thereby separating the liquid L in the storagecontainer 57 into the first liquid L1 and the second liquid L2.

Furthermore, in the above embodiment, the filter 60 is caused to traveldownward from the upper side of the liquid level of the liquid L in thestorage container 57 (the mixed liquid of the biological sample and thediluted solution) into the liquid to thereby separate the liquid L tothe liquid L1 and the liquid L2. However, another configuration also maybe used where the filter 60 is fixed at a predetermined position in thestorage container 57 and the liquid L is allowed to pass through thefilter 60 to thereby separate the liquid L into the liquid L1 and theliquid L2. For example, a configuration also may be used where thefilter 60 is fixed at a predetermined position at the lower side in thestorage container 57 and the liquid L in an amount for increasing theliquid level to the upper part of the filter 60 is introduced from thelower side of the filter 60 into the storage container 57 to therebyallow the liquid L to pass through the filter 60 to subsequently acquirethe liquid existing at the lower side of the filter 60 in the storagecontainer 57. In this case, the specimen quantitation section 27 as wellas the valves V1, V7, and V4 configure a liquid separation section forsending the liquid L through the filter 60 to separate the liquid L intothe liquid L1 and the liquid L2. This configuration also can allow onlycells other than an epidermal cell as a measurement target cell (e.g.,red blood cells and white blood cells) to pass through the filter 60 andthe epidermal cell as a measurement target cell is not allowed to passthrough the filter 60 and remains in the liquid at the lower side of thefilter 60 in the storage container 57. Thus, such liquid can be acquiredthat has a reduced number of cells other than the measurement targetcell.

Furthermore, in the above embodiment, a measurement sample prepared bythe sample preparation apparatus 3 is measured by a flow cytometer.However, another configuration also may be used that includes: a smearpreparation apparatus for smearing a measurement sample prepared by thesample preparation apparatus 3 to a glass slide to prepare a smear; anda cell image processing apparatus for imaging the prepared smear toanalyze an epidermal cell in the imaged image. Since a measurementsample for which the number of cells such as red blood cells or whiteblood cells is reduced is smeared to the glass slide, an epidermal cellas a measurement target cell can be analyzed accurately.

Furthermore, in the above embodiment, the liquid in the living bodycontainer 53 (mixed liquid of a biological sample and preservativesolution including methanol as a major component) is introduced to thestorage container 57 of the discrimination/substitution section 29 andthe diluted solution is subsequently supplied into the storage container57 and is subjected to a discrimination/substitution processing, therebysubstituting the preservative solution including methanol as a majorcomponent with the diluted solution. However, another configuration alsomay be used where solvent suitable for the PI staining for preparing ameasurement sample is substituted with preservative solution includingmethanol as a major component. As a result, a measurement sample can beprepared more favorably.

Furthermore, in the above embodiment, the storage container 57 is causedto be stationary and the filtration cylinder 58 is allowed to travel inthe up-and-down direction to thereby separate the liquid L in thestorage container 57 into the first liquid L1 and the second liquid L2.However, the present invention is not limited to this. Anotherconfiguration also may be used where the filtration cylinder 58 iscaused to be stationary and the storage container 57 is allowed totravel in the up-and-down direction to thereby separate the liquid L inthe storage container 57 into the first liquid L1 and the second liquidL2. Another configuration also may be used where, in synchronizationwith the travel of the filtration cylinder 58 in the up-and-downdirection, the storage container 57 is allowed to travel in the oppositedirection to thereby separate the liquid L in the storage container 57into the first liquid L1 and the second liquid L2.

Furthermore, in the above embodiment, the stirrer bar 68 provided in thestorage container 57 is rotated to thereby remove the first cell(epidermal cell) C1 attached to the lower face of the filter 60 by ashearing force. However, another configuration also may be used wherethe filtration cylinder 58 including the filter 60 is oscillated in thehorizontal direction or the filtration cylinder 58 is rotated in thehorizontal direction to thereby remove the first cell C1 attached to thelower face of the filter 60.

Furthermore, in the above embodiment, the rotation of the stirrer bar 68is followed by the application of a pressure from the upper side of thefilter 60 to the through hole of the filter 60. However, anotherconfiguration also may be used where the application of a pressure fromthe upper side of the filter 60 to the through hole of the filter 60 isfollowed by the rotation of the stirrer bar 68. Furthermore, in theabove embodiment, a pressure is applied from the upper side of thefilter 60 to the through hole of the filter 60 while the stirrer bar 68is being rotated. However, another configuration also may be used wherea pressure is applied to the through hole of the filter 60 while therotation of the stirrer bar 68 is being stopped.

Furthermore, in the above embodiment, the stoppage of the rotation ofthe stirrer bar 68 is followed by the sending of the concentrated liquidL1 in the storage container 57 to the outside of the storage container57. However, another configuration also may be used where theconcentrated liquid L1 is sent to the outside of the storage container57 while the stirrer bar 68 is being rotated.

We claim:
 1. A sample processing apparatus comprising: a samplecontainer configured to contain a sample solution including a biologicalsample collected from a cervix and a preservation solution, thebiological sample including epidermal cells of the cervix, red bloodcells, white blood cells and bacteria; a cell disperser configured todisperse the epidermal cells included in the sample solution containedin the sample container; a filtering unit which includes a first room, asecond room and a filter provided between the first room and the secondroom, wherein the filter is configured to prevent the epidermal cellsfrom passing therethrough and allow the red blood cells, the white bloodcells and the bacteria to pass therethrough; a sample supplierconfigured to supply, into the first room, the sample solution containedin the sample container; a pressure generator configured to generate apressure differential between the first room and the second room suchthat at least the preservation solution, the red blood cells, the whiteblood cells and the bacteria are moved from the first room to the secondroom via the filter; a substitution liquid supplier configured tosupply, into the first room, a substitution liquid which substitutes forthe preservation solution in the first room; and a stirrer that isarranged within the first room and is configured to rotate so as to movethe epidermal cells attached to the filter into the substitution liquidin the first room.
 2. The sample processing apparatus according to claim1, wherein the pressure generator comprises a negative pressure sourceconfigured to apply a negative pressure into the second room.
 3. Thesample processing apparatus according to claim 2, wherein the negativepressure source discharges at least the preservation solution, the redblood cells, the white blood cells and the bacteria which have beenmoved into the second room via the filter.
 4. The sample processingapparatus according to claim 3, further comprising a controllerconfigured to control the substitution liquid supplier and the negativepressure source so as to repeat the supply of the substitution liquidand the discharge of at least the preservation solution, the red bloodcells, the white blood cells and the bacteria.
 5. The sample processingapparatus according to claim 4, wherein the controller is configured tocontrol the substitution liquid supplier and the negative pressuresource so as to repeat the supply of the substitution liquid and thedischarge of at least the preservation solution, the red blood cells,the white blood cells and the bacteria until the preservation solutionwithin the first room is replaced with the substitution liquid.
 6. Thesample processing apparatus according to claim 1, wherein the samplesupplier comprises a sample flow path for supplying the sample solutioninto the first room.
 7. The sample processing apparatus according toclaim 1, wherein the substitution liquid supplier comprises asubstitution liquid flow path for supplying the substitution liquid intothe first room.
 8. The sample processing apparatus according to claim 1,further comprising a positive pressure source configured to apply apositive pressure into the second room so as to move the epidermal cellsattached to the filter into the substitution liquid in the first room.9. The sample processing apparatus according to claim 1, wherein thestirrer is configured to rotate in a direction along a filtering area ofthe filter.
 10. The sample processing apparatus according to claim 1,wherein a distance between the filtering area of the filter and thestirrer opposed to the filtering area is 1mm or less.
 11. The sampleprocessing apparatus according to claim 1, wherein the filter has athrough hole having a diameter of 8 to 20 μm.
 12. The sample processingapparatus according to claim 1, wherein the filtering unit has a concaveon a surface which forms the first room and faces the filter, and thestirrer is arranged within the concave.
 13. A sample preparing apparatuscomprising: a sample container configured to contain a sample solutionincluding a biological sample collected from a cervix and a preservationsolution, the biological sample including epidermal cells of the cervix,red blood cells, white blood cells and bacteria; a cell disperserconfigured to disperse the epidermal cells included in the samplesolution contained in the sample container; a filtering unit whichincludes a first room, a second room and a filter provided between thefirst room and the second room, wherein the filter is configured toprevent the epidermal cell from passing therethrough and allow the redblood cells, the white blood cells and the bacteria to passtherethrough; a sample supplier configured to supply, into the firstroom, the sample solution contained in the sample storage container; apressure generator configured to generate a pressure differentialbetween the first room and the second room such that at least thepreservation solution, the red blood cells, the white blood cells andthe bacteria are moved from the first room to the second room via thefilter; a substitution liquid supplier configured to supply, into thefirst room, a substitution liquid which substitutes for the preservationsolution in the first room; a stirrer that is arranged within the firstroom and is configured to rotate so as to move the epidermal cellsattached to the filter into the substitution liquid in the first room;and a reagent supplier configured to supply a reagent to a liquid takenout from the first room so as to prepare a measurement sample, whereinthe liquid taken out from the first room includes the substitutionliquid and the epidermal cells.
 14. The sample preparing apparatusaccording to claim 13, wherein the reagent is a staining solution forstaining the epidermal cells.
 15. The sample preparing apparatusaccording to claim 14, wherein the preservation solution containsalcohol.
 16. The sample preparing apparatus according to claim 13,wherein the pressure generator comprises a negative pressure sourceconfigured to apply a negative pressure into the second room.
 17. Thesample preparing apparatus according to claim 16, wherein the negativepressure source discharges at least the preservation solution, the redblood cells, the white blood cells and the bacteria which have beenmoved into the second room via the filter.
 18. The sample preparingapparatus according to claim 17, further comprising a controllerconfigured to control the substitution liquid supplier and the negativepressure source so as to repeat the supply of the substitution liquidand the discharge of at least the preservation solution, the red bloodcells, the white blood cells and the bacteria.
 19. The sample preparingapparatus according to claim 18, wherein the controller is configured tocontrol the substitution liquid supplier and the negative pressuresource so as to repeat the supply of the substitution liquid and thedischarge of the at least the preservation solution, the red bloodcells, the white blood cells and the bacteria until the preservationsolution within the first room is replaced with the substitution liquid.